Aspartic proteases are the targets for structure-based drug design for their role in physiological processes and pharmaceutical applications. Structural insights into the thermal inactivation mechanism of an aspartic protease in presence and absence of bound pepstatin A have been obtained by kinetics of thermal inactivation, CD, fluorescence spectroscopy and molecular dynamic simulations. The irreversible thermal inactivation of the aspartic protease comprised of loss of tertiary and secondary structures succeeded by the loss of activity, autolysis and aggregation The enthalpy and entropy of thermal inactivation of the enzyme in presence of pepstatin A increased from 81.2 to 148.5 kcal mol⁻¹, and from 179 to 359 kcal mol⁻¹ K⁻¹ respectively. Pepstatin A shifted the mid-point of thermal inactivation of the protease from 58 °C to 77 °C. The association constant (K) for pepstatin A with aspartic protease was 2.5 ± 0.3 × 10 ⁵ M⁻¹ and ΔG^o value was −8.3 kcal mol⁻¹. Molecular dynamic simulation studies were able to delineate the role of pepstatin A in stabilizing backbone conformation and side chain interactions. In the Cα-backbone, the short helical segments and the conserved glycines were part of the most unstable segments of the protein. Understanding the mechanism of thermal inactivation has the potential to develop re-engineered thermostable proteases.

天冬氨酸蛋白酶因其在生理过程和药物应用中的作用而成为基于结构的药物设计的目标。通过热灭活动力学、CD、荧光光谱和分子动力学模拟，已经获得了对存在和不存在结合胃蛋白酶抑制剂 A 的天冬氨酸蛋白酶的热灭活机制的结构见解。天冬氨酸蛋白酶的不可逆热失活包括三级和二级结构的丧失，继之以活性丧失、自溶和聚集。在胃蛋白酶抑制素 A 存在下，酶的热失活的焓和熵从 81.2 kcal mol ^-1增加到 148.5 kcal mol ^-1，和从 179 到 359 kcal mol ^-1  K ^-1分别。胃蛋白酶抑制剂 A 将蛋白酶的热灭活中点从 58 °C 移至 77 °C。胃酶抑素 A 与天冬氨酸蛋白酶的结合常数 ( K ) 为 2.5 ± 0.3 × 10 ⁵  M ^-1，Δ G ^o值为 -8.3 kcal mol ^-1。分子动力学模拟研究能够描述胃酶抑素 A 在稳定骨架构象和侧链相互作用中的作用。在 Cα 主链中，短螺旋片段和保守的甘氨酸是蛋白质最不稳定片段的一部分。了解热失活的机制有可能开发出重新设计的热稳定蛋白酶。